Unusual Acyltransferase Negatively Regulates Nigericin Production via Malonylation of P450 NigD_Sy

Secondary metabolism in Streptomyces is subjected to complex regulation. Compared to regulations on the transcriptional level, our knowledge of those on the post-translational level remains limited. Here, we identify a Gcn5-related N-acetyltransferase, SyPmatA, in the marine-derived Streptomyces youssoufiensis OUC6819, which negatively affects the production of nigericin, a polyether ionophore antibiotic with antifungal and antiparasitic activities. Inactivation of syPmatA enhanced the nigericin yield by 5.2-fold. Using in vitro and in vivo assays, we demonstrate that SyPmatA specifically catalyzes malonylation of P450 NigD_Sy at K224, negatively modulating its function. Overexpression of nigD_Sy^K224R (mimicking demalonylation) in the ΔsyPmatA mutant further increased nigericin production. Fermentation optimization and scale-up in a 5L-bioreactor using the engineered strain ΔsyPmatA/nigD_sy^K224R achieved a final titer of 444.5 mg/L, representing a 61.9-fold improvement over the wild-type strain. Our findings reveal a novel lysine malonylation-mediated regulatory mechanism in nigericin biosynthesis and highlight the potential of targeting post-translational modification enzymes for metabolic engineering.